The glycineamide ribonucleotide transformylase enzyme (chicken liver) has been purified to apparent homogeneity employing affinity chromatography. Copurifying with the enzyme is a second protein, formylmethenyl-methylene-tetrahydrofolate synthetase which suggests that the proteins are either covalently linked or specifically aggregated in the native state. We propose to (1) determine the stereospecificity of the transformylase towards the tetrahydrofolate cofactor since earlier work indicated an unusual lack of cofactor stereospecificity, (2) to find out whether the synthetase enzyme system is functional in synthesizing the cofactor required by the GAR transformylase enzyme; that is, starting with formic acid, ATP and tetrahydrofolic acid, whether a formylated GAR is produced as well as the relative efficiency of this process compared to commencing with the 5,10-methenyl tetrahydrofolate cofactor, (a similar set of experiments will be done starting with 5,10-methylene tetrahydrofolate and utilizing the dehydrogenase function of the combined enzyme), (3) to test whether the 5,11-methenyl derivative of tetrahydrohomofolate, an antimetabolite, will serve to inhibit the activity of either the combined enzyme or the GAR transformylase, and (4) to determine whether the transfer of the formyl group proceeds through a formylated enzyme intermediate.